Work

Multiplex quantitative SILAC for analysis of archaeal proteomes: a case study of oxidative stress responses

Year: 2017

Type: article

Abstract: Summary Stable isotope labelling of amino acids in cell culture (SILAC) is a quantitative proteomic method that can illuminate new pathways used by cells to adapt to different lifestyles and niches. Archaea, while thriving in extreme environments and accounting for ∼20%–40% of the Earth's biomass, have not been analyzed with the full potential of SILAC. Here, we report SILAC for quantitative comparison of archaeal proteomes, using Haloferax volcanii as a model. A double auxotroph was generated that allowed for complete incorporation of 13 C/ 15 N‐lysine and 13 C‐arginine such that each peptide derived from trypsin digestion was labelled. This strain was found amenable to multiplex SILAC by case study of responses to oxidative stress by hypochlorite. A total of 2565 proteins was identified by LC‐MS/MS analysis ( q ‐value ≤ 0.01) that accounted for 64% of the theoretical proteome. Of these, 176 proteins were altered at least 1.5‐fold ( p ‐value < 0.05) in abundance during hypochlorite stress. Many of the differential proteins were of unknown function. Those of known function included transcription factor homologs related to oxidative stress by 3D‐homology modelling and orthologous group comparisons. Thus, SILAC is found to be an ideal method for quantitative proteomics of archaea that holds promise to unravel gene function.

Source: Environmental Microbiology

DOI: 10.1111/1462-2920.14014

Authors: Lana J. McMillan, Sungmin Hwang, Rawan E. Farah, Jin Koh, Sixue Chen, Julie A. Maupin‐Furlow

Institutions: University of Florida, Institute of Food and Agricultural Sciences

Language: English

Topic: Advanced Proteomics Techniques and Applications

Subfield: Spectroscopy

Field: Chemistry

Domain: Physical Sciences

Open Access status: green